A mapping technique for probing the structure of proteoglycan core molecules.
Our previous work showed that treatment of chick embryo cartilage proteoglycan (PG-H) with chondroitinase-AC II and keratanase yielded a protein-rich core fraction having enzymatically modified linkage oligosaccharides. The core sample has now been analyzed by tryptic peptide mapping, in which the isolated core sample contained in a single Coomassie blue-staining band from a dried slab gel is radioiodinated and treated with trypsin, and the resultant tryptic peptides are displayed two-dimensionally on a silica gel thin layer plate. The map thus obtained exhibited 22 major peptide spots, the resolution and location of which were reproducible. In order to identify regions of the core polypeptide from which the tryptic peptides are derived, PG-H was cleaved with clostripain under conditions that yield a hyaluronic acid-binding fragment with an apparent Mr = 150,000 and chondroitin sulfate-peptide clusters of smaller molecular sizes. Although the peptide maps of the two size classes of clostripain fragments differed significantly from each other, the patterns of spots, as a whole, were extensively similar to those observed with the intact core molecule. These results have provided additional evidence that PG-H has a single, nonvariable core protein structure. In addition, the technique used here will provide a versatile method for the identification of genetic types in this increasingly complex family of matrix macromolecules.
D000818:Animals; D002356:Cartilage / Q000032:analysis*; D055598:Chemical Phenomena; D002621:Chemistry; D002642:Chick Embryo; D002808:Chondroitin Lyases; D002845:Chromatography; D003546:Cysteine Endopeptidases; D004586:Electrophoresis; D010450:Endopeptidases; D006026:Glycoside Hydrolases; D010446:Peptide Fragments; D010455:Peptides / Q000302:isolation & purification*; D011509:Proteoglycans / Q000032:analysis*; D014357:Trypsin; D001616:beta-Galactosidase
D010446:Peptide Fragments; D010455:Peptides; D011509:Proteoglycans; D006026:Glycoside Hydrolases; C026093:keratan-sulfate endo-1,4-beta-galactosidase; D001616:beta-Galactosidase; D010450:Endopeptidases; D014357:Trypsin; D003546:Cysteine Endopeptidases; C019913:clostripain; D002808:Chondroitin Lyases